Freeze-substitution of rabbit tibial articular cartilage reveals that radial zone collagen fibres are tubules

Investigations of the micromorphology of rabbit tibial articular cartilage using scanning and transmission electron microscopy revealed that the colla genous elements in the tissue form fluid-containing tubular structures. The commonly described radial or deep zone longitudinal fibres were found to b e tubular structures with internal diameters of 1-2 mu m. The walls of the tubules were composed of tightly packed fibrils of collagen, The tangential zone, close to the tibial plateau, was composed mainly of a spongy arrange ment of collagen fibrils, containing bunches of tangentially lying small (< 1 mu m) diameter tubules, The application of conventional chemical fixation techniques resulted in the fine detail of this tissue being obscured. When the tissue was frozen, followed by cryo-scanning electron microscopy or fr eeze-drying, prior to observation in the scanning electron microscope the t ubule structures Were not obviously present. It was only by applying freeze -substitution techniques, followed by critical point drying or resin embedd ing, that the structure was revealed dearly. Segregation of water into ice crystals did occur during the freezing process, but the formation of those crystals played no part in creating the tubular morphology observed, A simi lar structure was still revealed following pre-treatment with glycerol, met hanol or Triton X-100, provided that concentration of these additives was n ot too high. The walls of the tubules in the radial region were composed of straight, longitudinally arranged as well as helically arranged, 30 nm dia meter fibrils. The lumen of the tubules appears to be lined by a circumfere ntially arranged array of approximately 10 nm diameter fibres, spaced at re gular intervals of 50-70 nm.